Pluripotent stem cells in the bone marrow (bm) give rise to hematopoietic lineages that populate the adult. We observed an alteration on the developmental profile of marked decrease in cells with B-specific surface markers (B220 and BP-1)in the bm of aged mice when compared to young. Other hemopoietic lineages were only slightly changed in the aged bm. The decrease found in pre-B/immature B cell populations progresses with age and is detected as early as 12 mos. of age. Initially, antibodies specific for markers of the B cell stages (TdT, B220 and cytoplasmic IgM) coupled with mitotic arrest will be used to determine the turnover rate for cells in each compartment of the aged. If a particular step of development is altered, we will then focus on that stage of B cell development. Also the rate of reconstitution of B cells in the spleen after irradiation of mice whose long bones are shielded will be determined. Reconstitution of young lethally irradiated mice with aged bm gives a profile which is now strikingly young-like. These results will be repeated with donor cells hearing Ly5 allotypic markers. Antibodies will be used to positively identify the donor cells. This suggests that the microenvironment of the bm is responsible for the B cell age-related differences. Stromal cell lines (SCL) established from both young and aged bm will be tested for their support of B cell development. B lineages will be determined using the same markers which identified stages of B cells in the bm. The SCL from aged and young mice that differ in their ability to support B cell development will be supplemented with cytokines known to affect B cells and reanalyzed. Also, RNAs from these SCL will be studied using a novel PCR approach to determine transcriptional differences. The differences will permit the identification of genes that are regulated differentially in the SCL from the aged. With the changes altering the B cell developmental profile in the aged mouse, the anti-phosphorylcholine response in BALB/c mice will be determined in B cells from the bm and spleen using a plaque forming cell assay (PFC). Others have shown that this response in aged is quantitatively and qualitatively different in young mice. The PFC will be picked by micromanipulation, CDNA will be made and amplified so that expressed V genes can be subcloned for sequencing. Results of this project will define the V genes seen in the immune response in animals of different ages.